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园艺学报 ›› 2009, Vol. 36 ›› Issue (8): 1215-1220.

• 研究简报 • 上一篇    下一篇

扁桃AcSFB基因的克隆与原核表达分析

郭长奎1;罗淑萍1;李疆2*;高启明2   

  1. 郭长奎1;罗淑萍1;李疆2*;高启明2
  • 收稿日期:2009-02-24 修回日期:2009-07-02 出版日期:2009-08-25 发布日期:2009-08-25
  • 通讯作者: 李疆

Identification and Prokaryotic Expression of Almond AcSFB Gene

GUO Chang-kui1;LUO Shu-ping1;LI Jiang2*;GAO Qi-ming   

  1. (1College of Agronomy, Xinjiang Agricultural University, Urumqi 830052, China; 2College of Forestry and Horticulture, Xinjiang Agricultural University, Urumqi 830052, China)
  • Received:2009-02-24 Revised:2009-07-02 Online:2009-08-25 Published:2009-08-25
  • Contact: LI Jiang2

摘要: 利用RT-PCR和RACE技术, 从新疆栽培扁桃‘鹰嘴’花药中获得了一个全长的SFB ( S-hap-
lotype-specific F-box gene) 基因(GenBank登录号为FJ362525) , 命名为AcSFBd。该基因全长1 125 bp, 编码374个氨基酸, 在N末端含有一个大约50个氨基酸组成的F2box基序, 含有两个高变区(Hva和Hvb)和两个可变区(V1和V2) 。生物信息学的方法成功预测了AcSFBd蛋白分子质量为43 kD, 为亲水性、非分泌型蛋白, 在基质内起连接酶(EC6. - . - . -) 的作用。成功构建了原核表达载体, 并转化大肠杆菌BL21 (DE3) , SDS2PAGE检测结果显示表达了一个约60 kD的蛋白。

关键词: 扁桃, 基因, 克隆, SFB 基因, 原核表达, 生物信息学

Abstract: Style S-determinants have been determined to be S-RNase and the SFB ( S-haplotype-specific F-box gene) alleles are considered to be the first candidates of the pollen S-determinants. A total length SFB gene (GenBank accession No1FJ362525) named AcSFBd was cloned with RT-PCR and RACE technolgy from the anther of almond Yingzui p lanted in Xinjiang. AcSFBd was 1 125 bp in length encoding a protein of 374 amino acids. This gene encoded proteins with the F-box motif in the N amino terminus, and with four ( hyper) variable regions, i e. , V1, V2, Hva and Hvb which were similarwith S-RNase genes, considered to be recognition site between SFB and S-RNase. With the bioinformatics analysis, the AcSFBd protein was 43 kD,was instable, hydrophilicity and non-secretory protein and had ligase enzyme activity. After transformed to Escherichia coli BL21 and induced by isopropyl-β-D-thiogalactopyranoside ( IPTG) , recombinant protein about
60 kD was expressed in pET-AcSFB system and separated by SDS-PAGE electrophoresis

Key words: almond, gene, clone, SFB gene, p rokaryotic exp ression, bioinformatics

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